ATP is the main cellular energy generated by the enzyme ATP synthase in almost all organisms from bacteria to vertebrates. While malfunction of the ATP synthase complex is responsible for several disease conditions, the enzyme itself can be used as a potent molecular drug target to combat many diseases including microbial infections, cancer, tuberculosis, and obesity. Recent widespread escalation of antibiotic resistant microbes in general and E. coli in particular demands novel alternative approaches to combat microbial infections. Inhibition of ATP synthase by inhibitors such as peptides is known to deprive microbes of required energy, resulting in microbial cell death. Therefore, we have examined the venom peptide induced inhibition of E. coli ATP synthase. It was found that venom peptides completely inhibited E. coli ATP synthase and the process of inhibition was found to be fully reversible. This study also links the antimicrobial properties of peptides in part to the inhibition of ATP synthase. Thus, selective use of ATP synthase as a molecular drug may have an important impact on biology and medicine.